Neisseria are Gram-negative cocci, 0.6 to 1.0 μm in diameter. Two species of Neisseria are of major medical importance: N. gonorrhoeae, the causative agent of gonorrhoea, and N. meningitidis, the causative agent of bacterial meningitis. Humans are the only known reservoir of the members of the genus Neisseria. It is well recognized that for a vaccine against N. meningitidis or N. gonorrhoeae to be efficacious, bactericidal antibodies able to kill the bacterium in the presence of complement must be elicited. Current vaccine formulations against meningococcal B strains, for example, focus on polysaccharides and outer membrane proteins, which are highly variable proteins with little or no cross-reactivity between strains. In addition, there is no current vaccine for gonococcal strains. Accordingly, an improved immunogenic composition for both Neisseria gonorrhoeae and Neisseria meningitidis, which may be used in vaccine compositions, is highly desired. In addition, an immunogenic composition comprising an adjuvant which is able to elicit an antibactericidal response to N. meningitidis and N. gonorrhoeae is also highly desired.
Adjuvants are compounds which are capable of potentiating an immune response to antigens. Adjuvants can potentiate both humoral and cellular immunity. For certain pathogens, however, it is preferable to stimulate cellular immunity and, indeed, T cells. Presently used adjuvants do not adequately induce T cell responses, and/or have deleterious side effects.
Currently, the only adjuvants approved for human use in the United States are aluminum salts (alum). These adjuvants have been useful for some vaccines including hepatitis B, diphtheria, polio, rabies, and influenza, but may not be useful for others, especially if stimulation of cell-mediated immunity is required for protection. For example, reports indicate that alum failed to improve the effectiveness of whooping cough and typhoid vaccines and provided only a slight effect with adenovirus vaccines. Additionally, problems such as, induction of granulomas at the injection site and lot-to-lot variation of alum preparations have been experienced.
Complete Freund's adjuvant (CFA) is a powerful immunostimulatory agent that has been used successfully with many antigens on an experimental basis, and especially for animal research. CFA is comprised of three components: a mineral oil, an emulsifying agent such as Arlacel A, and killed mycobacteria such as Mycobacterium tuberculosis. Aqueous antigen solutions are mixed with these components to create a water-in-oil emulsion. CFA causes severe side effects, however, including pain, abscess formation, and fever, which prevent its use in either human or veterinary vaccines. The side effects are primarily due to the host's reactions to the mycobacterial component of CFA. Incomplete Freund's adjuvant (IFA) is similar to CFA without the bacterial component. While not yet approved for use in the United States, IFA has been useful for several types of vaccines in other countries. IFA has been used successfully in humans with influenza and polio vaccines and with several animal vaccines including rabies, canine distemper, and foot-and-mouth disease. Experiments have shown, however, that both the oil and emulsifier used in IFA can cause tumors in mice, indicating that an alternative adjuvant would be a better choice for human use.
Muramyl dipeptide (MDP) represents the minimal unit of the mycobacterial cell wall complex that generates the adjuvant activity observed with CFA. Ellouz et al., Biochem. Biophys. Res. Comm., 1974, 59, 1317. Many synthetic analogs of MDP have been generated that exhibit a wide range of adjuvant potency and side effects. Chedid et al., Prog. Allergy, 1978, 25, 63. Three analogs of MDP—threonyl derivatives of MDP (Byars et al.; Vaccine, 1987, 5, 223); n-butyl derivatives of MDP (Chedid et al., Infect. Immun., 1982, 35, 417); and lipophilic derivatives of muramyl tripeptide (Gisler et al., Immunomodulations of Microbial Products and Related Synthetic Compounds, Y. Yamamura and S. Kotani, Eds., Excerpta Medica, Amsterdam, p. 167)—have been shown to stimulate humoral and cell-mediated immunity and exhibit low levels of toxicity. Another derivative of MDP, N-acetylmuramyl-L-alanyl-D-isoglutaminyl-L-alanine-2-[1,2-d]palmitoyl-sn-glycero-3-3(hydroxyphosphoryl-oxy)]ethylamide (MTP-PE) is lipophilic. MTP-PE has phospholipid tails that allow association of the hydrophobic portion of the molecule with a lipid environment while the muramyl peptide portion associates with the aqueous environment. Thus, MTP-PE itself can act as an emulsifying agent to generate stable oil-in-water emulsions.
Levamisole and isoprinosine are other synthetic adjuvants that increase host immunity. Levamisole is the levo isomer of tetramisole and potentiates humoral and cellular immunity through a T cell-dependent mechanism. Isoprinosine, a complex containing inosine, the purine precursor of adenosine and guanosine, promotes T cell mitogenesis. Tuftsin, a 4 amino acid peptide (Thr-Lys-Pro-Arg) homologous to a sequence in the immunoglobulin (Ig) heavy chain, primarily stimulates macrophages.
Microparticles prepared from the biodegradable and biocompatible polymers, known as the poly(lactide-co-glycolides) (PLG), have been demonstrated to be effective adjuvants for a number of antigens. In addition, PLG microparticles can control the rate of release of entrapped antigens and, thus, offer potential for single-dose vaccines. Moreover, administration of biodegradable polymers with entrapped antigens have been demonstrated in a range of animal models to induce potent immune responses. O'Hagan et al., Advanced Drug Deliv. Rev., 1998, 32, 225-246 and Singh et al., Advanced Drug Deliv. Rev., 1998, 34, 285-304.
An emulsion comprising squalene, sorbitan trioleate (Span85™), and polysorbate 80 (Tween 80™) microfluidized to provide uniformly sized microdroplets, i.e. MF59, has also been shown to induce potent immune responses. MF59 formulations have been shown to induce antibody titers 5->100 times greater than those obtained with aluminum salt adjuvants. MF59 has been demonstrated to enhance the immune response to antigens from numerous sources including, for example, herpes simplex virus (HSV), human immunodeficiency virus (HIV), influenza virus, hepatitis C virus (HCV), cytomegalovirus (CMV), hepatitis B virus (HBV), human papillomavirus (HPV), and malaria. Ott et al., Vaccine Design: The Subunit And Adjuvant Approach, 1995, M. F. Powell and M. J. Newman, Eds., Plenum Press, New York, p. 277-296; Singh et al., Vaccine, 1998, 16, 1822-1827; Ott et al., Vaccine, 1995, 13, 1557-1562; O'Hagan et al., Mol. Medicine. Today, 1997, February, 69-75; and Traquina et al., J. Infect. Dis., 1996, 174, 1168-75, the disclosures of which are incorporated herein by reference in their entirety. MF59 adjuvant improves the immunogenicity of subunit antigens while maintaining the safety and tolerability profile of alum adjuvant. Van Nest et al., Vaccines 92, 1992, Cold Spring Harbor Laboratory Press, 57-62 and Valensi et al., J. Immunol., 1994, 153, 4029-39, the disclosures of which are incorporated herein by reference in their entirety. MF59 is further described in U.S. patent application Ser. No. 08/434,512, filed May 4, 1995, the disclosure of which is incorporated herein by reference in its entirety. In animal studies, MF59 has not been found to be genotoxic, teratogenic, nor cause sensitization. The mechanism of action of MF59 appears to be dependent upon the generation of a strong CD4+ T cell, i.e., a Th2 cell response. MF59 adjuvants, however, elicit little, if any, Th1 responses, or cytotoxic T lymphocyte (CTL) responses.
Oligonucleotides comprising CpG motifs mixed with antigens have been demonstrated to induce strong Th1 immune responses. Roman et al., Nat. Med., 1997, 3, 849-854; Weiner et al., Proc. Natl. Acad. Sci. USA, 1997, 94, 10833-10837; Davis et al., J. Immunol., 1998, 160, 870-876; Chu et al., J. Exp. Med., 1997, 186, 1623-1631; Lipford et al., Eur. J. Immunol., 1997, 27, 2340-2344; and Moldoveanu et al., Vaccine, 1988, 16, 1216-1224, the disclosures of which are incorporated herein by reference in their entirety. Unmethylated CpG dinucleotides are relatively common in bacterial DNA, but are underrepresented and methylated in vertebrate DNA. Bird, Trends Genet., 1987, 3, 342-347. Bacterial DNA or synthetic oligonucleotides containing unmethylated CpG motifs are also known to induce immune responses including, for example, B cell proliferation, interleukin-6 and immunoglobulin secretion, and apoptosis resistance. Krieg et al., Nature, 1995, 374, 546-549; Klinman et al., Proc. Natl. Acad. Sci. USA, 1996, 93, 2879-2883; Ballas et al., J. Immunol., 1996, 157, 1840-1845; Cowdery et al., J. Immunol., 1996, 156, 4570-4575; Halpern et al., Cell. Immunol., 1996, 167, 72-78; Yamamoto et al., Jpn. J. Cancer Res., 1988, 79, 866-873; Stacey et al., J. Immunol., 1996, 157, 2116-2122; Messina et al., J. Immunol., 1991, 147, 1759-1764; Yi et al., J. Immunol., 1996, 157, 4918-4925; Yi et al., J. Immunol., 1996, 157, 5394-5402; Yi et al., J. Immunol., 1998, 160, 4755-4761; and Yi et al., J. Immunol., 1998, 160, 5898-5906; PCT publications WO96/02555, WO98/16247, WO98/18810, WO98/40100, WO98/55495, WO98/37919 and WO98/52581, the disclosures of which are incorporated herein by reference in their entirety. CpG oligonucleotides, however, have not been shown to induce bactericidal antibody responses.
An effective immunogenic composition comprising an adjuvant in combination with a Neisseria antigen which induces bactericidal antibodies and which can be used for prophylactic and therapeutic treatment in animals for research is highly desired. In addition, an effective immunogenic composition comprising an adjuvant in combination with a Neisseria antigen which induces bactericidal antibodies and which can be used for prophylactic and therapeutic treatment in humans is also still desired. Such a response would be helpful in treatment of, for example, meningococcal or gonococcal infections as well as for immunizing individuals susceptible to meningococcal or gonococcal infections.